Dicarboxylic acids (diacids) are important compounds that are used in the manufacture of commercial polymers (e.g. polyesters, polyurethanes). The diacid adipic acid [1] is used mainly as a monomer in the production of 6,6-nylon, a polyamide generated through the reaction of [1] with hexane-1,6-diamine. Polyesters (for use in fabrics and plastics of many compositions) are formed through the polymerization of terephthalic acid [3] and a dialcohol (diol) such as ethylene glycol (to make polyethylene terephthalate), propane diol (poly(1,3-propanediol terephthalate)) or butanediol (poly(1,4-butanediolphthalate). Adipic acid is also used in the synthesis of various polyesters. Currently adipic acid is synthesized via oxidation of cyclohexane and similar petrochemicals using traditional chemical synthesis.
The large scale worldwide use of nylons and polyesters requires the production of millions of metric tons of [1] and [3] annually. These diacids are themselves synthesized from starting materials extracted from petroleum. One means of reducing the large dependence on oil for the commercial production of polymers is to generate the diacids by a fermentation process involving the use of polyketide synthases.
The use of hybrid polyketide synthases to produce diacids with a carbon backbone with an odd number of carbon atoms is disclosed in International Patent Application No. PCT/US2009/038831, filed Mar. 30, 2009, which published as PCT publication no. WO 2009/121066 on Oct. 1, 2009. The use of hybrid polyketide synthases to produce diacids is disclosed in U.S. Patent Application Pub. No. 2013/0280766, now issued as U.S. Pat. No. 9,334,514.
The polyketides are one of the most diverse and chemically complicated classes of molecules known, its members frequently weighing in excess of 500 daltons and harboring numerous stereocenters. Partly owing to their antibacterial, immunosuppressive, and anti-cancer activities, much effort has been devoted to deciphering the mechanism by which polyketide synthases (PKSs) synthesize their products. PKSs perform Claisen condensation reactions between a loaded acyl-ACP intermediate and an α-substituted (H, CH3, C2H5, etc.) malonyl-CoA extender unit analogous to fatty acid biosynthesis. This is then followed by varying degrees of 3-reduction by accessory domains. This condensation-reduction cycle is repeated by subsequent downstream modules until the intermediate is liberated from the enzyme, most commonly by the activity of a thioesterase domain (reviewed in (Khosla, 2009)).
Engineering of type I modular PKSs has the potential to produce an enormous variety of novel, rationally-designed compounds. Yet, more than two decades after their modular nature was discovered (Donadio et al., 1991), there are currently no commercial applications of engineered PKSs.